High-fat High-cholesterol Diet Research Articles

Impact of Atorvastatin on Skeletal Muscle Mitochondrial Activity, Locomotion and Axonal Excitability-Evidence from ApoE-/- Mice.

The cardiovascular benefit of statins is well established. However, only 20% of high-risk patients remain adequately adherent after 5 years of treatment. Among reasons for discontinuation, statin associated-muscle pain symptoms are the most prevalent. Aim of the present study was to evaluate the impact of high dose atorvastatin on skeletal muscle mitochondrial activity, aerobic and anaerobic exercise, and axonal excitability in a murine model of atherosclerosis. ApoE-/- mice were fed 12 weeks a high-fat high-cholesterol diet alone or containing atorvastatin (40 mg/Kg/day). Outcomes were the evaluation of muscle mitochondrial functionality, locomotion, grip test, and axonal excitability (compound action potential recording analysis of Aα motor propioceptive, Aβ mechanoceptive and C nociceptive fibres). Atorvastatin led to a reduction in muscle mitochondrial biogenesis and mitochondrial ATP production. It did not affect muscular strength but led to a time-dependent motor impairment. Atorvastatin altered the responsiveness of mechanoceptive and nociceptive fibres, respectively, the Aβ and C fibres. These findings point out to a mild sensitization on mechanical, tactile and pain sensitivity. In conclusion, although the prevalence of muscular side effects from statins may be overestimated, understanding of the underlying mechanisms can help improve the therapeutic approach and reassure adherence in patients needing-to-be-treated.

Genetic Deficiency of Indoleamine 2,3-dioxygenase Aggravates Vascular but Not Liver Disease in a Nonalcoholic Steatohepatitis and Atherosclerosis Comorbidity Model

Nonalcoholic steatohepatitis (NASH) is a chronic liver disease that increases cardiovascular disease risk. Indoleamine 2,3-dioxygenase-1 (IDO1)-mediated tryptophan (Trp) metabolism has been proposed to play an immunomodulatory role in several diseases. The potential of IDO1 to be a link between NASH and cardiovascular disease has never been investigated. Using Apoe−/− and Apoe−/−Ido1−/− mice that were fed a high-fat, high-cholesterol diet (HFCD) to simultaneously induce NASH and atherosclerosis, we found that Ido1 deficiency significantly accelerated atherosclerosis after 7 weeks. Surprisingly, Apoe−/−Ido1−/− mice did not present a more aggressive NASH phenotype, including hepatic lipid deposition, release of liver enzymes, and histopathological parameters. As expected, a lower L-kynurenine/Trp (Kyn/Trp) ratio was found in the plasma and arteries of Apoe−/−Ido1−/− mice compared to controls. However, no difference in the hepatic Kyn/Trp ratio was found between the groups. Hepatic transcript analyses revealed that HFCD induced a temporal increase in tryptophan 2,3-dioxygenase (Tdo2) mRNA, indicating an alternative manner to maintain Trp degradation during NASH development in both Apoe−/− and Apoe−/−Ido1−/mice−. Using HepG2 hepatoma cell and THP1 macrophage cultures, we found that iron, TDO2, and Trp degradation may act as important mediators of cross-communication between hepatocytes and macrophages regulating liver inflammation. In conclusion, we show that Ido1 deficiency aggravates atherosclerosis, but not liver disease, in a newly established NASH and atherosclerosis comorbidity model. Our data indicate that the overexpression of TDO2 is an important mechanism that helps in balancing the kynurenine pathway and inflammation in the liver, but not in the artery wall, which likely determined disease outcome in these two target tissues.

Abstract 153: The Differential Effects Of Weight Loss And Regain On Atherosclerosis

Heightened inflammation in obesity is a major contributor to many diseases, including atherosclerosis and other cardiovascular diseases (CVD). Although weight loss is extremely beneficial in improving many obesity-related conditions, weight regain worsens disease outcomes, including CVD, compared to maintenance of an obese state. Nonetheless, how weight loss promotes inflammation resolution and why weight regain worsens inflammation is poorly understood and was investigated here. Using high-fat high-cholesterol diet feeding of atherogenic mice (Ldlr deficient), we promoted weight loss after the establishment of atherosclerosis and obesity by using dietary restriction, e.g. reducing daily food consumption by 30% for 2 weeks, while maintaining hyperlipidemia. For weight regain, some mice were then given free access to food for 6 weeks. Strikingly, after 2 weeks of dietary restriction we observed atherosclerosis resolution, while weight regain accelerated disease re-progression. Single-cell analysis of visceral adipose and plaque leukocytes revealed enrichment in both tissues of a novel macrophage population with dietary restriction, which is distinguished by high expression of the antibody receptor Fcgr4, a known phagocytosis mediator. Conversely, Fcgr4 + macrophages disappeared from plaques and adipose following weight regain. Mechanistically, Fcgr4 + macrophages from the adipose tissue induced clearance of plaque necrotic core via enhanced efferocytosis. Moreover, we discovered that weight loss and regain differentially influence immune progenitors. Bone marrow (BM) transplantation showed that while weight loss and obese BM recipients had similar plaque inflammation, weight regain BM recipients had larger, more inflammatory plaques. In conclusion, our data are the first to directly show that initial weight loss promotes resolution of atherosclerosis, independently of plasma cholesterol changes and through the enrichment of specialized pro-resolving macrophages in adipose tissue that communicate with plaques. Conversely, weight regain accelerates disease progression via inflammatory reprogramming of immune progenitors.

Cholesterol Promotes Colorectal Cancer Growth by Activating the PI3K/AKT Pathway.

Globally, the incidence of colorectal cancer (CRC) increases each year, with an unhealthy diet representing one of the major pathogenic risk factors for CRC. Cholesterol is a vital dietary ingredient required to maintain the normal function of the body; however, disturbances in cholesterol levels have been discovered to exert a significant role in tumorigenesis. The present study is aimed at investigating the role of cholesterol in the occurrence of CRC. Briefly, CRC model mice were established through an intraperitoneal injection of azoxyemethane (AOM) and were subsequently either fed a normal diet (ND), high-fat diet (HFD), or high-fat high-cholesterol diet (HFHC). Furthermore, in vitro experiments were performed following the treatment of SW480 and HCT116 cells with cholesterol, and the cell viability and colony formation rate of CRC cells were analyzed. The findings identified that cholesterol levels were increased in CRC tissues compared with adjacent normal tissues. In contrast, the serum levels of cholesterol were decreased in patients with CRC compared with the healthy controls; however, no significant differences were observed in the cholesterol levels between stage I + II and stage III + IV patients with CRC. Notably, CRC model mice fed with an HFD or HFHC recorded a larger body weight compared with those mice fed a ND; however, no significant differences were reported in the number of tumors formed in each group. Furthermore, the tumor size in the HFHC group was discovered to be increased compared with the ND and HFD groups, and HGD and the pathological morphology were the most pronounced in the HFHC group. Moreover, mice in the HFHC group presented the highest ratio of Ki-67-positive staining and the lowest ratio of TUNEL-positive staining compared with those in the two other groups. Cholesterol treatment also increased the cell viability and clonality of SW480 and HCT116 cells. In addition, the protein expression levels of phosphorylated-AKT were upregulated in cholesterol-induced CRC cells and tissues, whereas the treatment with BAY80-6946 attenuated the cholesterol-induced increases in the cell viability, colony formation ability, and tumor size. In conclusion, the findings of the present study suggested that cholesterol may stimulate the progression of CRC by activating the PI3K/AKT signaling pathway; however, cholesterol may not affect the number of tumors formed in CRC. In addition, cholesterol was discovered to mainly affect the advanced stages of CRC rather than the early stages.

Receptor-interacting protein kinase-1 ablation in liver parenchymal cells promotes liver fibrosis in murine NASH without affecting other symptoms.

Non-alcoholic steatohepatitis (NASH), a chronic liver disease that emerged in industrialized countries, can further progress into liver fibrosis, cirrhosis, and hepatocellular carcinoma. In the next decade, NASH is predicted to become the leading cause of liver transplantation, the only current interventional therapeutic option. Hepatocyte death, triggered by different death ligands, plays key role in its progression. Previously, we showed that the receptor-interacting protein kinase-1 (RIPK1) in hepatocytes exhibits a protective role in ligand-induced death. Now, to decipher the role of RIPK1 in NASH, Ripk1LPC-KO mice, deficient for RIPK1 only in liver parenchymal cells, and their wild-type littermates (Ripk1fl/fl) were fed for 3, 5, or 12weeks with high-fat high-cholesterol diet (HFHCD). The main clinical signs of NASH were analyzed to compare the pathophysiological state established in mice. Most of the symptoms evolved similarly whatever the genotype, whether it was the increase in liver to body weight ratio, the steatosis grade or the worsening of liver damage revealed by serum transaminase levels. In parallel, inflammation markers followed the same kinetics with significant equivalent inductions of cytokines (hepatic mRNA levels and blood cytokine concentrations) and a main peak of hepatic infiltration of immune cells at 3weeks of HFHCD. Despite this identical inflammatory response, more hepatic fibrosis was significantly evidenced at week 12 in Ripk1LPC-KO mice. This coincided with over-induced rates of transcripts of genes implied in fibrosis development (Tgfb1, Tgfbi, Timp1, and Timp2) in Ripk1LPC-KO animals. In conclusion, our results show that RIPK1 in hepatocyte limits the progression of liver fibrosis during NASH.

Label-free Assessment of the Nascent State of Rat Non-alcoholic Fatty Liver Disease Using Spontaneous Raman Microscopy.

Spontaneous Raman microscopy, which can detect molecular vibrations in cells and tissues, could be a useful tool for the label-free assessment of non-alcoholic fatty liver disease (NAFLD). However, it is unclear whether it can be used to evaluate the nascent state of NAFLD. To address this, we analyzed the Raman spectra of rat liver tissues in the nascent state of NAFLD upon excitation at 532 nm. Raman and histochemical analyses were performed of liver tissues from rats fed a high-fat, high-cholesterol diet (HFHCD). Raman microscopic imaging analysis of formalin-fixed thin tissue slices showed hepatic steatosis, as revealed by the Raman band at 2,854 cm−1, whereas lipid droplets were not detectable by hematoxylin-eosin staining of images until 3 days after feeding a HFHCD. Raman signals of retinol at 1,588 cm−1 emitted from hepatic stellate cells were distributed alongside hepatic cords; the retinol content rapidly decreased after feeding a HFHCD, whereas hepatic lipid content increased inversely. Raman microscopic analysis of the surface of fresh ex vivo livers enabled early detection of lipid accumulation after a 1-day feeding a HFHCD. In conclusion, spontaneous Raman microscopy can be applied to the label-free evaluation of the nascent state of NAFLD liver tissues.

Limosilactobacillus fermentum prevents gut-kidney oxidative damage and the rise in blood pressure in male rat offspring exposed to a maternal high-fat diet.

Oxidative stress along the gut-kidney axis is a risk factor for developing arterial hypertension in offspring from dams fed a high-fat diet. Considering the antioxidant capacity of probiotic strains, this study evaluated the effects of a daily multistrain formulation with Limosilactobacillus fermentum 139, 263, and 296 on blood pressure (BP), renal function, and oxidative stress and along the gut-kidney axis in male offspring from dams fed a high-fat high-cholesterol (HFHC) diet during pregnancy and lactation. Dams were fed a diet control or HFHC diet during pregnancy and lactation. At 100 days of age, part of the male offspring from dams fed a HFHC diet received Limosilactobacillus fermentum formulation for 4 weeks (HFHC + Lf) daily. After the 4-week intervention, BP (tail-cuff plethysmography) and urinary and biochemical variables were measured. In addition, malondialdehyde levels, enzymatic activities of superoxide dismutase, catalase, glutathione-S-transferase, and nonenzymatic antioxidant defense (thiols content) were measured in the colon and renal cortex. Male offspring from dams fed a HFHC had increased blood pressure, impaired renal function, and oxidative stress along the gut-kidney axis. Administration of Limosilactobacillus fermentum reduced systolic, diastolic, and mean blood pressure levels and alleviated renal function impairment and oxidative stress along the gut-kidney axis in male offspring from dams fed a HFHC diet. Administration of Limosilactobacillus fermentum formulation attenuated programmed hypertension in the HFHC group through oxidative stress modulation along the gut-kidney axis.

Lacticaseibacillus rhamnosus FM9 and Limosilactobacillus fermentum Y57 Are as Effective as Statins at Improving Blood Lipid Profile in High Cholesterol, High-Fat Diet Model in Male Wistar Rats.

Elevated serum cholesterol is a major risk factor for coronary heart diseases. Some Lactobacillus strains with cholesterol-lowering potential have been isolated from artisanal food products. The purpose of this study was to isolate probiotic Lactobacillus strains from traditional yoghurt (dahi) and yogurt milk (lassi) and investigate the impact of these strains on the blood lipid profile and anti-obesity effect in a high cholesterol high fat diet model in Wistar rats. Eight candidate probiotic strains were chosen based on in vitro probiotic features and cholesterol reduction ability. By 16S rDNA sequencing, these strains were identified as Limosilactibacillus fermentum FM6, L. fermentum FM16, L. fermentum FM12, Lacticaseibacillus rhamnosus FM9, L. fermentum Y55, L. fermentum Y57, L. rhamnosus Y59, and L. fermentum Y63. The safety of these strains was investigated by feeding 2 × 108 CFU/mL in saline water for 28 days in a Wistar rat model. No bacterial translocation or any other adverse effects were observed in animals after administration of strains in water, which indicates the safety of strains. The cholesterol-lowering profile of these probiotics was evaluated in male Wistar rats using a high-fat, high-cholesterol diet (HFCD) model. For 30 days, animals were fed probiotic strains in water with 2 × 108 CFU/mL/rat/day, in addition to a high fat, high cholesterol diet. The cholesterol-lowering effects of various probiotic strains were compared to those of statin. All strains showed improvement in total cholesterol, LDL, HDL, triglycerides, and weight gain. Serum cholesterol levels were reduced by 9% and 8% for L. rhamnosus FM9 and L. fermentum Y57, respectively, compared to 5% for the statin-treated group. HDL levels significantly improved by 46 and 44% for L. rhamnosus FM9 and L. fermentum Y57, respectively, compared to 46% for the statin-treated group. Compared to the statin-treated group, FM9 and Y57 significantly reduced LDL levels by almost twofold. These findings show that these strains can improve blood lipid profiles as effectively as statins in male Wistar rats. Furthermore, probiotic-fed groups helped weight control in animals on HFCD, indicating the possible anti-obesity potential of these strains. These strains can be used to develop food products and supplements to treat ischemic heart diseases and weight management. Clinical trials, however, are required to validate these findings.

Purple sweet potato anthocyanin extract regulates redox state related to gut microbiota homeostasis in obese mice.

This study explored the advantageous effects of purple sweet potato anthocyanin extract (PSPAE) on redox state in obese mice. The normal chow diet (NCD) group, high-fat/cholesterol diet (HCD) group, and three groups based on HCD and added with low, middle, and high dose of PSPAE (PAL, PAM, and PAH) were raised for 12weeks. High dose of PSPAE treatment decreased the elevations of the body weight by 24.7%, serum total cholesterol by 48.3%, serum triglyceride by 42.4%, and elevated serum activities of glutathione peroxidase by 53.3%, superoxide dismutase by 57.8%, catalase by 75.4%, decreased serum contents of malondialdehyde by 27.1% and lipopolysaccharides by 40.5%, as well as increased caecal total short-chain fatty acid by 2.05-fold. Additionally, PSPAE depressed toll-like receptor 4 (TLR-4), nuclear factor kappa-B (NF-κB), interleukin 6, tumor necrosis factor α, and preserved nuclear factor erythroid-2-related factor 2 (Nrf2) gene expression. Similarly, the protein expression of Nrf2 was enhanced, while TLR-4 and p-NF-κB/NF-κB were depressed by PSPAE treatment. Moreover, PSPAE administration promoted the protection of intestinal barrier function and rebuilt gut microbiota homeostasis by blooming g_Akkermansia, g_Bifidobacterium, and g_Lactobacillus. Furthermore, antibiotic interference experiments showed that the gut microbiota was indispensable for preserving the redox state of PSPAE. These results suggested that PSPAE administration could be an opportunity for improving HCD-induced obesity and the redox state related to gut dysbiosis. PRACTICAL APPLICATION: Purple sweet potato anthocyanin has diverse pharmacological properties. It is applicable for individuals to consume extracts (as pills or other forms) from raw purple sweet potato if they want to improve obesity or redox state.

Ganoderic acid A ameliorates non-alcoholic streatohepatitis (NASH) induced by high-fat high-cholesterol diet in mice.

Non-alcoholic steatohepatitis (NASH) is becoming a huge global health problem. Previous studies have revealed that ganoderic acids have hepatoprotective and hypocholesterolemic effects. In the present study, to evaluate the anti-NASH activity of ganoderic acid A (GAA), male 6-week-old C57BL/6J mice were divided into the following four groups, which were administered different diets: Normal diet (ND group), high-fat high-cholesterol diet (HFHC group), HFHC diet supplemented with 25 mg/kg/day (GAAL group) or 50 mg/kg/day of GAA (GAAH group). After 12 weeks of GAA treatment, histopathological results revealed that compared with that of the HFHC group, GAA significantly inhibited fat accumulation, steatosis, inflammation and fibrosis in the liver. GAA effectively reduced serum aspartate transaminase and alanine transaminase levels compared with the HFHC model. Furthermore, the endoplasmic reticulum (ER) stress-responsive proteins, including glucose-regulated protein 78, phosphorylated (p)-eukaryotic initiation factor-2α and p-JNK, were significantly suppressed by GAA, while ERp57, p-MAPK and p-AKT were significantly increased after GAA treatment. Taken together, it was concluded that GAA could resist HFHC diet-induced NASH. In terms of its underlying mechanism, GAA could improve liver inflammation and fibrosis by inhibiting hepatic oxidative stress and the ER stress response induced by HFHC.

A high-fat/high-cholesterol diet, but not high-cholesterol alone, increases free cholesterol and apoE-rich HDL serum levels in rats and upregulates hepatic ABCA1 expression

A high-fat/high-cholesterol (HFC) diet, but not a high-cholesterol (HC) diet, is known to induce elevated serum apolipoprotein E (apoE)-rich high-density lipoprotein (HDL) levels in animal models. However, the exact mechanisms by which the combination of dietary fat and cholesterol induces apoE-rich HDL production is not well understood. Here, we investigated the effects of dietary fat and cholesterol on serum lipoprotein profiles and hepatic mRNA expression that are associated with HDL production, cholesterol transport, and bile acid metabolism. Male Sprague-Dawley rats were fed HFC, HC, high-fat, or control diets and then evaluated. The HFC diet induced significant increases in hepatic free-cholesterol accumulation (1.4-fold, p < 0.01) and serum apoE-rich HDL cholesterol (8.7-fold, p < 0.001) levels compared with the HC diet. The apoE-rich HDL induced by the HFC diet was remarkably rich in free cholesterol. Liver gene-expression was mostly similar between the HC and HFC diet groups. However, there was a significant increase of ATP-binding cassette transporter A1 (ABCA1) levels in the HFC group compared to the HC group for both mRNA (1.9-fold, p < 0.001) and protein (6.6-fold, p < 0.01). These results suggest that an increase in apoE-rich HDL induced by dietary fat and cholesterol may be involved in cholesterol efflux from the liver through increased ABCA1-mediated free-cholesterol efflux.

Switching to Regular Diet Partially Resolves Liver Fibrosis Induced by High-Fat, High-Cholesterol Diet in Mice

The globally prevalent disease, non-alcoholic steatohepatitis (NASH), is characterized by a steatotic and inflammatory liver. In NASH patients, tissue repair mechanisms, activated by the presence of chronic liver damage, lead to the progressive onset of hepatic fibrosis. This scar symptom is a key prognostic risk factor for liver-related morbidity and mortality. Conflicting reports discuss the efficiency of dietary interventions on the reversibility of advanced fibrosis established during NASH. In the present study, the effect of dietary interventions was investigated in the outcome of the fibrosis settled in livers of C57BL/6J mice on a high-fat, high-cholesterol diet (HFHCD) for 5 or 12 consecutive weeks. Various clinico-pathological investigations, including a histological analysis of the liver, measurement of plasma transaminases, steatosis and fibrosis, were performed. To assess the effectiveness of the dietary intervention on established symptoms, diseased mice were returned to a standard diet (SD) for 4 or 12 weeks. This food management resulted in a drastic reduction in steatosis, liver injuries, inflammatory markers, hepatomegaly and oxidative stress and a gradual improvement in the fibrotic state of the liver tissue. In conclusion, our results demonstrated that dietary intervention can partially reverse liver fibrosis induced by HFHCD feeding.

Premorbid Steatohepatitis Increases the Seriousness of Dextran Sulfate Sodium-induced Ulcerative Colitis in Mice.

Background and AimsThe concurrence of nonalcoholic steatohepatitis (NASH) and ulcerative colitis (UC) is increasingly seen in clinical practice, but the underlying mechanisms remain unclear. This study aimed to develop a mouse model of the phenomenon by combining high-fat high-cholesterol diet (HFHCD)-induced NASH and dextran sulfate sodium (DSS)-induced UC, that would support mechanistic studies.MethodsMale C57BL/6 mice were randomly assigned to two groups receiving either a chow diet or HFHCD for 12 weeks of NASH modeling. The mice were the divided into four subgroups for UC modeling: (1) A control group given a chow diet with normal drinking water; (2) A colitis group given chow diet with 2% DSS in drinking water; (3) A steatohepatitis group given HFHCD with normal drinking water; and (4) A steatohepatitis + colitis group given HFHCD with 2% DSS in drinking water.ResultsNASH plus UC had high mortality (58.3%). Neither NASH nor UC alone were fatal. Although DSS-induced colitis did not exacerbate histological liver injury in HFHCD-fed mice, premorbid NASH significantly increased UC-related gut injury compared with UC alone. It was characterized by a significantly shorter colon, more colonic congestion, and a higher histopathological score (p<0.05). Inflammatory (tumor necrosis factor-alpha, interleukin 1 beta, C-C motif chemokine ligand 2, and nuclear factor kappa B) and apoptotic (Bcl2, Bad, Bim, and Bax) signaling pathways were significantly altered in distal colon tissues collected from mice with steatohepatitis + colitis compared with the other experimental groups.ConclusionsPremorbid steatohepatitis significantly aggravated DSS-induced colitis and brought about a lethal phenotype. Potential links between NASH and UC pathogeneses can be investigated using this model.

High Fat High Cholesterol (HFHC) Diet Alters Liver and Uterine Histopathology in Late Pregnant Rats

The aim of the present study was to reveal the effect of the high-fat high cholesterol diet (HFHC) in pregnant rats. HFHC diet was fed to Wistar female rats 8 weeks before and after mating upto 20th day of gestation. The effect of the HFHC diet was studied by measuring body weight, fat depots, liver weight, lipid profile, fetal parameters and histopathology of liver and uterus. HFHC diet fed rats showed significant increase in the fat depots, serum and uterine tissue cholesterol level. Increased liver weight, macrovesicular fatty degeneration of liver and infiltration of inflammatory cells and cystic endometrial glands were noticed in the uterine histopathology of HFHC fed rats. However, there was no change in litter size and weight. In conclusion, feeding of a HFHC diet prior to and during pregnancy of Wistar rats led to significant change in the metabolic profile of rats, which was associated with increase in liver weight and fat depots. Further, histopathology of liver and uterus showed inflammatory changes.

Nobiletin Prevents High-Fat Diet-Induced Dysregulation of Intestinal Lipid Metabolism and Attenuates Postprandial Lipemia.

Nobiletin is a dietary flavonoid that improves insulin resistance and atherosclerosis in mice with metabolic dysfunction. Dysregulation of intestinal lipoprotein metabolism contributes to atherogenesis. The objective of the study was to determine if nobiletin targets the intestine to improve metabolic dysregulation in both male and female mice. Approach and Results: Triglyceride-rich lipoprotein (TRL) secretion, intracellular triglyceride kinetics, and intestinal morphology were determined in male and female LDL (low-density lipoprotein) receptor knockout (Ldlr-/-), and male wild-type mice fed a standard laboratory diet or high-fat, high-cholesterol (HFHC) diet ± nobiletin using an olive oil gavage, radiotracers, and electron microscopy. Nobiletin attenuated postprandial TRL levels in plasma and enhanced TRL clearance. Nobiletin reduced fasting jejunal triglyceride accumulation through accelerated TRL secretion and lower jejunal fatty acid synthesis with no impact on fatty acid oxidation. Fasting-refeeding experiments revealed that nobiletin led to higher levels of phosphorylated AKT (protein kinase B) and FoxO1 (forkhead box O1) and normal Srebf1c expression indicating increased insulin sensitivity. Intestinal length and weight were diminished by HFHC feeding and restored by nobiletin. Both fasting and postprandial plasma GLP-1 (glucagon-like peptide-1; and likely GLP-2) were elevated in response to nobiletin. Treatment with a GLP-2 receptor antagonist, GLP-2(3-33), reduced villus length in HFHC-fed mice but did not impact TRL secretion in any diet group. In contrast to males, nobiletin did not improve postprandial lipid parameters in female mice. Nobiletin opposed the effects of the HFHC diet by normalizing intestinal de novo lipogenesis through improved insulin sensitivity. Nobiletin prevents postprandial lipemia because the enhanced TRL clearance more than compensates for increased TRL secretion.

Sphingosine 1-Phosphate Receptor 4 Promotes Nonalcoholic Steatohepatitis by Activating NLRP3 Inflammasome.

Background & AimsSphingosine 1-phosphate receptors (S1PRs) are a group of G-protein–coupled receptors that confer a broad range of functional effects in chronic inflammatory and metabolic diseases. S1PRs also may mediate the development of nonalcoholic steatohepatitis (NASH), but the specific subtypes involved and the mechanism of action are unclear.MethodsWe investigated which type of S1PR isoforms is activated in various murine models of NASH. The mechanism of action of S1PR4 was examined in hepatic macrophages isolated from high-fat, high-cholesterol diet (HFHCD)-fed mice. We developed a selective S1PR4 functional antagonist by screening the fingolimod (2-amino-2-[2-(4- n -octylphenyl)ethyl]-1,3- propanediol hydrochloride)-like sphingolipid-focused library.ResultsThe livers of various mouse models of NASH as well as hepatic macrophages showed high expression of S1pr4. Moreover, in a cohort of NASH patients, expression of S1PR4 was 6-fold higher than those of healthy controls. S1pr4+/- mice were protected from HFHCD-induced NASH and hepatic fibrosis without changes in steatosis. S1pr4 depletion in hepatic macrophages inhibited lipopolysaccharide-mediated Ca++ release and deactivated the Nod-like receptor pyrin domain-containning protein 3 (NLRP3) inflammasome. S1P increased the expression of S1pr4 in hepatic macrophages and activated NLRP3 inflammasome through inositol trisphosphate/inositol trisphosphate–receptor–dependent [Ca++] signaling. To further clarify the biological function of S1PR4, we developed SLB736, a novel selective functional antagonist of SIPR4. Similar to S1pr4+/- mice, administration of SLB736 to HFHCD-fed mice prevented the development of NASH and hepatic fibrosis, but not steatosis, by deactivating the NLRP3 inflammasome.ConclusionsS1PR4 may be a new therapeutic target for NASH that mediates the activation of NLRP3 inflammasome in hepatic macrophages.

Ursodeoxycholic Acid Treatment Restores Gut Microbiota and Alleviates Liver Inflammation in Non-Alcoholic Steatohepatitic Mouse Model.

Gut microbiota dysbiosis plays an important role in the progression of non-alcoholic fatty liver disease (NAFLD), and no approved drugs are available for NAFLD treatment. In this study, we aimed to explore the dynamic changes of gut microbiota at the different stages of NAFLD and determine whether ursodeoxycholic acid (UDCA) could improve liver histopathological features of non-alcoholic steatohepatitis (NASH) mice induced by a high-fat high-cholesterol (HFHC) diet and its impact on gut microbiota. 6-week-old male C57BL/6 mice were fed with a HFHC or normal diet for 12, 18, and 24 weeks, respectively, to simulate the different stages of NAFLD. 16s ribosomal RNA genes from mice fecal samples at the different time points were sequenced to evaluate the dynamic changes of the gut microbiota. Then, C57BL/6 mice were fed with a HFHC diet for 24 weeks to establish the NASH model. Different doses of UDCA were administered intragastrically for additional 4 weeks. Normal diet–fed mice were taken as control. Serum samples, liver, and intestine tissues were harvested for biochemical tests and histopathological examinations. 16s ribosomal RNA genes from mice fecal samples were sequenced to assess the structural changes of gut microbiota. HFHC diet–fed mice developed simple steatosis, steatohepatitis, and fibrosis at 12, 18, and 24 weeks, respectively. The profile of gut microbiota dynamically changed with the different stages of NAFLD. NASH mice had significantly higher abundance of Fecalibaculum, Coriobacteriaceae_UCG-002, and Enterorhabdus, and lower abundance of norank_f_Muribaculaceae, Bacteroides, and Alistipes, which were partially restored by UDCA treatment. UDCA treatment significantly attenuated hepatic inflammation of NASH mice as indicated by the sum of ballooning and lobular inflammation of the NALFD activity score (3.2 ± 0.8 vs 1.8 ± 0.8, p = 0.029), and partially restored gut microbiota dysbiosis, and increased the expression of Claudin-1 and ZO-1 in the intestine, but did not activate the suppressed Farnesoid X receptor signal pathway. Conclusions: The gut microbiota dynamically changes with the different stages of NAFLD. UDCA treatment (120 mg/kg) could partially restore gut microbiota, repair gut barrier integrity, and attenuate hepatic inflammation in the NASH mouse model.

Atorvastatin protects against liver and vascular damage in a model of diet induced steatohepatitis by resetting FXR and GPBAR1 signaling.

Farnesoid-x-receptor (FXR) agonists, currently trialed in patients with non-alcoholic steatosis (NAFLD), worsen the pro-atherogenic lipid profile and might require a comedication with statin. Here we report that mice feed a high fat/high cholesterol diet (HFD) are protected from developing a pro-atherogenic lipid profile because their ability to dispose cholesterol through bile acids. This protective mechanism is mediated by suppression of FXR signaling in the liver by muricholic acids (MCAs) generated in mice from chenodeoxycholic acid (CDCA). In contrast to CDCA, MCAs are FXR antagonists and promote a CYP7A1-dependent increase of bile acids synthesis. In mice feed a HFD, the treatment with obeticholic acid, a clinical stage FXR agonist, failed to improve the liver histopathology while reduced Cyp7a1 and Cyp8b1 genes expression and bile acids synthesis and excretion. In contrast, treating mice with atorvastatin mitigated liver and vascular injury caused by the HFD while increased the bile acids synthesis and excretion. Atorvastatin increased the percentage of 7α-dehydroxylase expressing bacteria in the intestine promoting the formation of deoxycholic acid and litocholic acid, two GPBAR1 agonists, along with the expression of GPBAR1-regulated genes in the white adipose tissue and colon. In conclusion, present results highlight the central role of bile acids in regulating lipid and cholesterol metabolism in response to atorvastatin and provide explanations for limited efficacy of FXR agonists in the treatment of NAFLD.

Achillea Fragrantissima Ethanolic Extract Exerts Hypocholesterolemia and Hepatic Antioxidant Effects in High Fat-Cholesterol Diet: An Experimental Study

Background: Hypercholesterolemia and oxidative stress consider the main causes for atherosclerotic cardiovascular diseases, that are one of the major non-communicable diseases responsible for more than a third of deaths in Saudi Arabia. Cholesterol-lowering medications as Atorvastatin® (ATOR) are linked to a variety of side effects. Achillea fragarntissima (AF) is a valuable medicinal plant in Saudi Arabia with potent antioxidant activity. Aim: The current study was performed to determine the efficacy of AF in the treatment of hypercholesterolemia through the antioxidant metabolic pathway. Methodology: Dried aerial parts of AF were extracted by ethanol (70%). Induction of hypercholesterolemia in rats was induced through feeding a high fat-cholesterol diet (HFCD) for 8 weeks. Rats were assigned to two main groups; control group (Cont, n=10) rats fed a standard diet, and hypercholesterolemic group (HFCD) (n=40) rats fed HFCD. The HFCD group was further assigned after measured lipid profile to confirm the induction of hypercholesterolemia to HFCD; HFCD+AF (hypercholesterolemic rats treated orally with 500 mg/kg AF); HFCD+ ATOR (hypercholesterolemic rats treated orally with 20 mg/kg ATOR, as a reference drug); and HFCD+AF+ATOR (hypercholesterolemic rats treated orally with AF+ ATOR). Different treatments were ingested to rats for 4 weeks. Results: The results revealed that the HFCD group showed significant hyperlipidemia (elevation of serum TC, TG, LDL-C, and VLDL-C levels concurrent with a reduction in serum HDL-C level); significant disturbance in liver functions (elevation in serum ALT, AST, and ALP enzymes activities); and significant oxidative stress (elevation in hepatic MDA level with a reduction in hepatic SOD activity) compared with the Cont group. Besides, hepatic central vein section showed deposition of large lipid within hepatocytes and abundant focal cell necrosis. Oral treatment with AF, ATOR, and the mixture of the drug and AF produced significant hypocholesterolemia, antioxidant, and improved liver function enzymes, with normalized hepatic central vein tissue compared with the HFCD group. The mixture of AF+ATOR had a superior effect than either treatment alone. Conclusion: In hypercholesterolemic rats, AF may be used to prevent atherosclerosis through improving lipid profile levels, protecting against hepatic oxidative stress, and ameliorating hepatic functions. Thus highlighting its valuable effects in the treatment of atherosclerotic cardiovascular diseases. Keywords: Achillea fragarntissima, lipid profile, hepatic oxidative stress, hepatic function, hypercholesterolemia.

Protection of CAPE-pNO2 Against Chronic Myocardial Ischemia by the TGF-Β1/Galectin-3 Pathway In Vivo and In Vitro.

Although it is known that caffeic acid phenethyl ester (CAPE) and its derivatives could ameliorate acute myocardial injury, their effects on chronic myocardial ischemia (CMI) were not reported. This study aimed to investigate the potential effect of caffeic acid p-nitro phenethyl ester (CAPE-pNO2, a derivative of CAPE) on CMI and underlying mechanisms. SD rats were subjected to high-fat-cholesterol-diet (HFCD) and vitamin D3, and the H9c2 cells were treated with LPS to establish CMI model, followed by the respective treatment with saline, CAPE, or CAPE-pNO2. In vivo, CAPE-pNO2 could reduce serum lipid levels and improve impaired cardiac function and morphological changes. Data of related assays indicated that CAPE-pNO2 downregulated the expression of transforming growth factor-β1 (TGF-β1) and galectin-3 (Gal-3). Besides, CAPE-pNO2 decreased collagen deposition, the number of apoptotic cardiomyocytes, and some related downstream proteins of Gal-3 in the CMI rats. Interestingly, the effects of CAPE-pNO2 on TGF-β1, Gal-3, and other proteins expressed in the lung were consistent with that in the heart. In vitro, CAPE-pNO2 could attenuate the fibrosis, apoptosis, and inflammation by activating TGF-β1/Gal-3 pathway in LPS-induced H9c2 cell. However, CAPE-pNO2-mediated cardioprotection can be eliminated when treated with modified citrus pectin (MCP, an inhibitor of Gal-3). And in comparison, CAPE-pNO2 presented stronger effects than CAPE. This study indicates that CAPE-pNO2 may ameliorate CMI by suppressing fibrosis, inflammation, and apoptosis via the TGF-β1/Gal-3 pathway in vivo and in vitro.